Molecular cloning and characterization of omega class glutathione S-transferase (GST-O) from the polychaete Neanthes succinea: biochemical comparison with theta class glutathione S-transferase (GST-T)

Comp Biochem Physiol C Toxicol Pharmacol. 2007 Nov;146(4):471-7. doi: 10.1016/j.cbpc.2007.05.003. Epub 2007 May 29.

Abstract

We cloned and sequenced a full-length cDNA of an omega class glutathione S-transferase (GST-O) from the polychaete Neanthes succinea (ns-GST-O). The full-length cDNA of ns-GST-O was 1562 bp in length, containing an open reading frame (OR) of 732 bp that encoded a 244 amino acid protein. The deduced amino acid sequence of ns-GST-O showed a low similarity with the theta class N. suucinea GST (ns-GST-T). As GSTs play a significant role in antioxidant defense, we checked the expression pattern of ns-GST-O in N. succinea after exposure to copper (CuCl(2) 12 to 72 mug/L), which is an oxidative stress-inducing agent. After exposure to CuCl(2), ns-GST-O gene was dramatically up-regulated and when compared with ns-GST-T the expression pattern was more pronounced at all the concentrations of copper. Even the basal transcription levels of ns-GST-O were higher than those of ns-GST-T. To further characterize the catalytic properties of ns-GST-O, we constructed a recombinant ns-GST-O plasmid with a 6x His-Tag at the N-terminal of the full-length ns-GST-O cDNA. Recombinant ns-GST-O protein was highly expressed in transformed Escherichia coli. The effect of pH, temperature and chemical inhibitors on the enzyme activity of ns-GST-O was also studied and compared with the reported effect of these factors on recombinant ns-GST-T protein. These results suggest that, like other types of GSTs, ns-GST-O protein plays a conserved antioxidant role in the polychaete N. succinea.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antioxidants / physiology
  • Base Sequence
  • Biomarkers
  • Cloning, Molecular
  • Copper / toxicity
  • Dose-Response Relationship, Drug
  • Gene Expression Regulation, Enzymologic*
  • Glutathione Transferase / classification
  • Glutathione Transferase / genetics*
  • Glutathione Transferase / metabolism*
  • Hydrogen-Ion Concentration
  • Molecular Sequence Data
  • Polychaeta / classification
  • Polychaeta / enzymology*
  • Polychaeta / genetics*
  • RNA, Messenger / metabolism
  • Recombinant Proteins
  • Water Pollutants, Chemical / toxicity

Substances

  • Antioxidants
  • Biomarkers
  • RNA, Messenger
  • Recombinant Proteins
  • Water Pollutants, Chemical
  • Copper
  • Glutathione Transferase
  • cupric chloride